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Si us plau utilitzeu sempre aquest identificador per citar o enllaçar aquest document: https://hdl.handle.net/2445/68711
Ultra-Sensitive Detection of Plasmodium falciparum by Amplification of Multi-Copy Subtelomeric Targets
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BACKGROUND: Planning and evaluating malaria control strategies
relies on accurate definition of parasite prevalence in the
population. A large proportion of asymptomatic parasite
infections can only be identified by surveillance with molecular
methods, yet these infections also contribute to onward
transmission to mosquitoes. The sensitivity of molecular
detection by PCR is limited by the abundance of the target
sequence in a DNA sample; thus, detection becomes imperfect at
low densities. We aimed to increase PCR diagnostic sensitivity
by targeting multi-copy genomic sequences for reliable detection
of low-density infections, and investigated the impact of these
PCR assays on community prevalence data. METHODS AND FINDINGS:
Two quantitative PCR (qPCR) assays were developed for
ultra-sensitive detection of Plasmodium falciparum, targeting
the high-copy telomere-associated repetitive element 2 (TARE-2,
approximately 250 copies/genome) and the var gene acidic
terminal sequence (varATS, 59 copies/genome). Our assays reached
a limit of detection of 0.03 to 0.15 parasites/mul blood and
were 10x more sensitive than standard 18S rRNA qPCR. In a
population cross-sectional study in Tanzania, 295/498 samples
tested positive using ultra-sensitive assays. Light microscopy
missed 169 infections (57%). 18S rRNA qPCR failed to identify 48
infections (16%), of which 40% carried gametocytes detected by
pfs25 quantitative reverse-transcription PCR. To judge the
suitability of the TARE-2 and varATS assays for high-throughput
screens, their performance was tested on sample pools. Both
ultra-sensitive assays correctly detected all pools containing
one low-density P. falciparum-positive sample, which went
undetected by 18S rRNA qPCR, among nine negatives. TARE-2 and
varATS qPCRs improve estimates of prevalence rates, yet other
infections might still remain undetected when absent in the
limited blood volume sampled. CONCLUSIONS: Measured malaria
prevalence in communities is largely determined by the
sensitivity of the diagnostic tool used. Even when applying
standard molecular diagnostics, prevalence in our study
population was underestimated by 8% compared to the new assays.
Our findings highlight the need for highly sensitive tools such
as TARE-2 and varATS qPCR in community surveillance and for
monitoring interventions to better describe malaria epidemiology
and inform malaria elimination efforts.
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HOFMANN, Natalie e., MWINGIRA, Felista, SHEKALAGHE, Seif, ROBINSON, Leanne j., MUELLER, Ivo, FELGER, Ingrid. Ultra-Sensitive Detection of Plasmodium falciparum by
Amplification of Multi-Copy Subtelomeric Targets. _PLoS Medicine_. 2015. Vol. 12, núm. 3, pàgs. e1001788. [consulta: 25 de febrer de 2026]. ISSN: 1549-1277. [Disponible a: https://hdl.handle.net/2445/68711]